Human tularemia is a highly virulent bacterial zoonosis with endemic foci in the northern hemisphere. Its clinical manifestations depend on the route of infection. The ulceroglandular form is the most common presentation after handling sources contaminated with Francisella tularensis. When ingested, contaminated food or water can cause an oropharyngeal form. Pulmonary, typhoidal (the two more common forms reported in laboratory workers), glandular and ocular forms are other less frequent presentations. The disease occurs in outbreaks, usually associated with direct contact with infected game or contaminated water, or in a seasonal pattern in arthropod-borne tularemia. The severity of this infection, its initial nonspecific manifestations, and the ability of the agent to survive in the environment have led to the inclusion of F. tularensis in a list of bacterial pathogens that could be used for bioterrorism. The human and murine responses to F. tularensis have been studied with particular emphasis on the survival of the bacterium within macrophages and the cytokine responses resulting from intracellular infection. Less known are the interactions of this organism with cells that it must encounter to cause systemic infection. Thus, a very focused study of the interaction of F. tularensis with endothelium is proposed for this R03 application. Both the attenuated vaccine strain and the virulent American strain (F. tularensis tularensis) will be used for parallel experiments on infection of endothelium and its pro-inflammatory activation, as measured by upregulation of expression of adhesion molecules and chemokines. One feature of tularemia that has not been investigated is the manner of spread of this organism within the infected host. Tularemic lesions contain a marked mononuclear cell infiltrate, and the sequence of lesion formation is not known. Trafficking of the bacterium alone and of cells infected with the bacterium (neutrophils, monocytes) will be studied by assessing their ability to cross endothelium in vitro. Earlier studies have found that the lipopolysaccharide of F. tularensis does not activate mononuclear cells, so its effect on stimulation of endothelial cells is not clear, nor is there a certainty that there is a CD14-dependency of the response to the lipopolysaccharide or the whole organism. The proposed research is aimed at a greater understanding of the mechanisms used by F. tularensis to cause systemic infection.